The present invention relates generally to organic chemistry, biochemistry, pharmacology and medicine. More particularly, it relates to novel heterocyclic compounds, and their physiologically acceptable salts and prodrugs, which modulate the activity of protein kinases (xe2x80x9cPKsxe2x80x9d) and, therefore, are expected to exhibit a salutary effect against disorders related to abnormal PK activity.
The following is offered as background information only and is not admitted to be or describe prior art to the present invention.
PKs are enzymes that catalyze the phosphorylation of hydroxy groups on tyrosine, serine and threonine residues of proteins. The consequences of this seemingly simple activity are staggering; cell growth, differentiation and proliferation; i.e., virtually all aspects of cell life in one way or another depend on PK activity. Furthermore, abnormal PK activity has been related to a host of disorders, ranging from relatively non-life threatening diseases such as psorisasis to extremely virulent diseases such as glioblastoma (brain cancer).
The PKs can conveniently be broken down into two classes, the protein tyrosine kinases (PTKs) and the serine-threonine kinases (STKs).
One of the prime aspects of PTK activity is its involvement with growth factor receptors. Growth factor receptors are cell-surface proteins. When bound by a growth factor ligand, growth factor receptors are converted to an active form which interacts with proteins on the inner surface of a cell membrane. This leads to phosphorylation on tyrosine residues of the receptor and other proteins and to the formation inside the cell of complexes with a variety of cytoplasmic signaling molecules that, in turn, affect numerous cellular responses such as cell division (proliferation), cell differentiation, cell growth, expression of metabolic effects to the extracellular microenvironment, etc. For a more complete discussion, see Schlessinger and Ullrich, Neuron, 9:303-391 (1992) which is incorporated by reference, including any drawings, as if fully set forth herein.
Growth factor receptors with PK activity are known as receptor tyrosine kinases (xe2x80x9cRTKsxe2x80x9d). They comprise a large family of transmembrane receptors with diverse biological activity. At present, at least nineteen (19) distinct subfamilies of RTKs have been identified. An example of these is the subfamily designated the xe2x80x9cHERxe2x80x9d RTKs, which include EGFR (epithelial growth factor receptor), HER2, HER3 and HER4. These RTKs consist of an extracellular glycosylated ligand binding domain, a transmembrane domain and an intracellular cytoplasmic catalytic domain that can phosphorylate tyrosine residues on proteins.
Another RTK subfamily consists of insulin receptor (IR), insulin-like growth factor I receptor (IGF-1R) and the insulin receptor related receptor (IRR). IR and IGF-1R interact with insulin, IGF-I and IGF-II to form a heterotetramer of two entirely extracellular glycosylated xcex1 subunits and two xcex2 subunits which cross the cell membrane and which contain the tyrosine kinase domain.
A third RTK subfamily is referred to as the platelet derived growth factor receptor (xe2x80x9cPDGFRxe2x80x9d) group, which includes PDGFRxcex1, PDGFRxcex2, CSFIR, c-kit and c-fms. These receptors consist of glycosylated extracellular domains composed of variable numbers of immunoglobin-like loops and an intracellular domain wherein the tyrosine kinase domains is interrupted by unrelated amino acid sequences.
Another group which, because of its similarity to the PDGFR subfamily, is sometimes subsumed in the later group is the fetus liver kinase (xe2x80x9cflkxe2x80x9d) receptor subfamily. This group is believed to be made of up of kinase insert domain-receptor fetal liver kinase-1 (KDR/FLK-1), flk-1R, flk-4 and fms-like tyrosine kinase 1 (fit-1).
One further member of the tyrosine kinase growth factor receptor family is the fibroblast growth factor (xe2x80x9cFGFxe2x80x9d) receptor group. This groups consists of four receptors, FGFR1-4, and seven ligands, FGF1-7. While not yet well defined, it appears that the receptors consist of a glycosylated extracellular domain containing a variable number of immunoglobin-like loops and an intracellular domain in which the PTK sequence is interrupted by regions of unrelated amino acid sequences.
A more complete listing of the known RTK subfamilies is described in Plowman et al., DNandP, 7(6):334-339 (1994) which is incorporated by reference, including any drawings, as if fully set forth herein.
In addition to the RTKs, there also exists a family of entirely intracellular PTKs called xe2x80x9cnon-receptor tyrosine kinasesxe2x80x9d or xe2x80x9ccellular tyrosine kinasesxe2x80x9d. This latter designation, abbreviated xe2x80x9cCTKxe2x80x9d, will be used herein. CTKs do not contain extracellular and transmembrane domains. At present, over 24 CTKs in 11 subfamilies (Src, Frk, Btk, Csk, Abl, Zap70, Fes/Fps, Fak, Jak and Ack) have been identified. The Src subfamily appear so far to be the largest group of CTKs and includes Src, Yes, Fyn, Lyn, Lck, Blk, Hck, Fgr and Yrk. For a more detailed discussion of CTKs, see Bolen, Oncogene, 8:2025-2031 (1993), which is incorporated by reference, including any drawings, as if fully set forth herein.
The serine-threonine kinases or STKS, like the CTKs, are predominantly intracellular although there are a few receptor kinases of the STK type. STKs are the most common of the cytosolic kinases; i.e., kinases which perform their function in that part of the cytoplasm other than the cytoplasmic organelles and cytoskelton. The cytosol is the region within the cell where much of the cell""s intermediary metabolic and biosynthetic activity occurs; e.g., it is in the cytosol that proteins are synthesized on ribosomes.
RTKs, CTKs and STKs have all been implicated in a host of pathogenic conditions including, significantly, large number of diverse cancers. Others pathogenic conditions which have been associated with PTKs include, without limitation, psoriasis, hepatic cirrhosis, diabetes, atherosclerosis, angiogenesis, restinosis, ocular diseases, rheumatoid arthritis and other inflammatory disorders, autoimmune disease and a variety of renal disorders.
With regard to cancer, two of the major hypotheses advanced to explain the excessive cellular proliferation that drives tumor development relate to functions known to be PK regulated. That is, it has been suggested that malignant cell growth results from a breakdown in the mechanisms that control cell division and/or differentiation. It has been shown that the protein products of a number of proto-oncogenes are involved in the signal transduction pathways that regulate cell growth and differentiation. These protein products of proto-oncogenes include the extracellular growth factors, transmembrane growth factor PTK receptors (RTKs), cytoplasmic PTKs (CTKs) and cytosolic STKs, discussed above.
In view of the apparent link between PK-related cellular activities and a number of human disorders, it is no surprise that a great deal of effort is being expended in an attempt to identify ways to modulate PK activity. Some of these have involved biomimetic approaches using large molecules patterned on those involved in the actual cellular processes (e.g., mutant ligands (U.S. App. Ser. No. 4,966,849); soluble receptors and antibodies (App. No. WO 94/10202, Kendall and Thomas, Proc. Nat""l Acad. Sci., 90:10705-09 (1994), Kim, et al., Nature, 362:841-844 (1993)); RNA ligands (Jelinek, et al., Biochemistry, 33:10450-56); Takano, et al., Mol. Bio. Cell 4:358A (1993); Kinsella, et al., Exp. Cell Res. 199:56-62 (1992); Wright, et al., J. Cellular Phys., 152:448-57) and tyrosine kinase inhibitors (WO 94/03427; WO 92/21660; WO 91/15495; WO 94/14808; U.S. Pat. No. 5,330,992; Mariani, et al., Proc. Am. Assoc. Cancer Res., 35:2268 (1994)).
More recently, attempts have been made to identify small molecules which act as PK inhibitors. For example, bis-monocylic, bicyclic and heterocyclic aryl compounds (PCT WO 92/20642), vinylene-azaindole derivatives (PCT WO 94/14808) and 1-cyclopropyl-4-pyridylquinolones (U.S. Pat. No. 5,330,992) have been described as PTK inhibitors. Styryl compounds (U.S. Pat. No. 5,217,999), styryl-substituted pyridyl compounds (U.S. Pat. No. 5,302,606), quinazoline derivatives (EP App. No. 0 566 266 A1), selenaindoles and selenides (PCT WO 94/03427), tricyclic polyhydroxylic compounds (PCT WO 92/21660) and benzylphosphonic acid compounds (PCT WO 91/15495) have all been described as PTK inhibitors useful in the treatment of cancer.
Our own efforts to identify small organic molecules which modulate PK activity and which, therefore, should be useful in the treatment and prevention of disorders driven by abnormal PK activity, has led us to the discovery of a family of novel 3-heteroarylidene-2-indolinone derivatives which exhibit excellent PK modulating ability and which are the subject of this invention.
The present invention relates generally to novel 3-heteroarylidene-2-indolinones which modulate the activity of receptor protein tyrosine kinases (RTK), non-receptor protein tyrosine kinases (CTKs) and serine/threonine protein kinases (STK). In addition, the present invention relates to the preparation and use of pharmacological compositions of the disclosed compounds and their physiologically acceptable salts and prodrugs in the treatment or prevention of PK driven disorders such as, by way of example and not limitation, cancer, diabetes, hepatic cirrhosis, atherosclerosis, angogenesis and renal disease.
A xe2x80x9c3-heteroarylidene-2-indolinonexe2x80x9d refers to a chemical compound having the general structure shown in Formula 1.
A xe2x80x9cpharmacological compositionxe2x80x9d refers to a mixture of one or more of the compounds described herein, or physiologically acceptable salts or prodrugs thereof, with other chemical components, such as physiologically acceptable carriers and excipients. The purpose of a pharmacological composition is to facilitate administration of a compound to an organism.
A xe2x80x9cprodrugxe2x80x9d refers to an agent which is converted into the parent drug in vivo. Prodrugs are often useful because, in some situations, they may be easier to administer than the parent drug. They may, for instance, be bioavailable by oral administration whereas the parent drug is not. The prodrug may also have improved solubility in pharmacological compositions over the parent drug. An example, without limitation, of a prodrug would be a compound of the present invention wherein it is administered as an ester (the xe2x80x9cprodrugxe2x80x9d) to facilitate transmittal across a cell membrane where water solubility is not beneficial, but then it is metabolically hydrolyzed to the carboxylic acid once inside the cell where water solubility is beneficial.
As used herein, an xe2x80x9cesterxe2x80x9d is a carboxyl group, as defined herein, wherein Rxe2x80x3 is any of the listed groups other than hydrogen.
As used herein, a xe2x80x9cphysiologically acceptable carrierxe2x80x9d refers to a carrier or diluent that does not cause significant irritation to an organism and does not abrogate the biological activity and properties of the administered compound.
An xe2x80x9cexcipientxe2x80x9d refers to an inert substance added to a pharmacological composition to further facilitate administration of a compound. Examples, without limitation, of excipients include calcium carbonate, calcium phosphate, various sugars and types of starch, cellulose derivatives, gelatin, vegetable oils and polyethylene glycols.
A. General Structural Features.
In one aspect, the compounds of the present invention relate to 3-heteroarylidene-2-indolinones which, in addition to being otherwise optionally substituted on both the heteroaryl and 2-indolinone portions of the compound, are necessarily substituted on the heteroaryl moiety (that is, on xe2x80x9cQxe2x80x9d of FIG. 1) with one or more hydrocarbon chains which themselves are substituted on the carbon furthest from the heteroaryl ring with a polar group. Physiologically acceptable salts and prodrugs of the claimed compounds are also within the scope of this invention.
A xe2x80x9chydrocarbon chainxe2x80x9d refers to an alkyl, alkenyl or alkynyl group, as defined herein.
A xe2x80x9cpolarxe2x80x9d group refers to a group wherein the nuclei of the atoms covalently bound to each other to form the group do not share the electrons of the covalent bond(s) joining them equally; that is the electron cloud is denser about one atom than another. This results in one end of the covalent bond(s) being relatively negative and the other end relatively positive; i.e., there is a negative pole and a positive pole. Examples of polar groups include, without limitation, hydroxy, alkoxy, carboxy, nitro, cyano, amino, quaternary ammonium, amido, ureido, sulfonamido, sulfinyl, sulfonyl, phosphono, morpholino, piperazinyl and tetrazolo.
While not being bound to any particular theory, applicants at this time believe that the polar groups may interact electronically, for example, but without limitation, through hydrogen bonding, Van der Walls forces and ionic bonds (but not covalent bonding), with the amino acids of a PTK active site. These interactions may assist the molecules of this invention to bind to the active site and thus prevent the natural substrate from entering the site. These polar groups may also contribute to the selectivity of some compounds; i.e., a particular polar group may have greater affinity for some PTK binding domains over others so that the compound is more potent against the former.
The terms xe2x80x9c2-indolinonexe2x80x9d and xe2x80x9c2-oxindolexe2x80x9d are used interchangeably herein; both refer to a chemical compound having the general structure: 
wherein A, B, D and E are carbon. It is to be understood, however, that this invention also features compounds wherein A, B, D and/or E are nitrogen; thus, wherever the terms xe2x80x9c2-indolinonexe2x80x9d or xe2x80x9c2-oxindolexe2x80x9d are used herein, they are to be construed as including the nitrogen analogs as well.
Thus, in another aspect, the present invention relates to compounds having the following chemical structure: 
A, B, D and E are selected from the group consisting of carbon and nitrogen, it being understood that the nitrogen-containing 9-member bicyclic ring formed is one known in the chemical arts; it being further understood that when A, B, D, or E is nitrogen, R3, R4, R5 or R6, respectively, does not exist;
R1 is selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, hydroxy, alkoxy, carboxyl, C-amido and sulfonyl.
R2 is selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, heteroaryl, and heteroalicyclic.
R3, R4, R5 and R6 are independently selected from the group consisting of hydrogen, alkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, sulfonamido, carbonyl, carboxyl, C-amido, N-amido, cyano, nitro, halo, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, amino and xe2x80x94NR10R11.
R10 and R11 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl, sulfonyl and, combined, a five- or six-member heteroalicyclic ring containing at least one nitrogen;
R3 and R4, R4 and R5, or R4 and R5 may combine to form a six-member aryl or heteroaryl ring.
Q is a heteroaryl group having the following structure: 
J is selected from the group consisting of oxygen, nitrogen and sulfur.
K, L and M are independently selected from the group consisting of carbon, nitrogen, oxygen and sulfur such that the five-member heteroaryl ring formed is one known in the chemical arts. It is understood that when K, L or M is nitrogen, sulfur or oxygen, R8 or -(alk1)nZ cannot be covalently bonded to that atom.
When J is nitrogen, R7 is selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, carbonyl, C-amido, guanyl, carboxyl, sulfonyl and trihalomethanesulfonyl and when J is oxygen or sulfur, R7 does not exist.
R8 is selected from the group consisting of hydrogen, alkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, sulfonamido, carbonyl, carboxyl, cyano, nitro, halo, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, amino, xe2x80x94NR10R11, trihalomethyl, a five member cycloalkyl, aryl, heteroaryl or heteoalicyclic ring fused to two adjacent atoms of the Q ring; and a six-member cycloalky, aryl, heteroaryl, or heteroalicyclic ring fused to two adjacent atoms of the Q ring.
R10 and R11 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl, sulfonyl and, combined, a five- or six-member heteroalicyclic ring containing at least one nitrogen.
Alk1 is selected from the group consisting of optionally substituted methylene (xe2x80x94CRRxe2x80x2xe2x80x94), optionally substituted ethylene (xe2x80x94C(R)xe2x95x90C(Rxe2x80x2)xe2x80x94) and acetylene(xe2x80x94Cxe2x89xa1Cxe2x80x94).
R and Rxe2x80x2 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, alkoxy, thioalkoxy, aryloxy and halo.
As for n, it is 1 to 10, inclusive.
Z is a polar group.
Examples of bicyclic groups known in the chemical arts include, but are not limited to:
Examples of xe2x80x9cfive atom heteroaryl groups known in the chemical artxe2x80x9d include, but are not limited to: 
As used herein, the term xe2x80x9calkylxe2x80x9d refers to a saturated aliphatic hydrocarbon including straight chain and branched chain groups. Preferably, the alkyl group has 1 to 20 carbon atoms (whenever it appears herein, a numerical range such as xe2x80x9c1 to 20xe2x80x9d refers to each integer in the given range; e.g., xe2x80x9c1 to 20 carbon atomsxe2x80x9d means that the alkyl group may consist of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc. up to and including 20 carbon atoms). More preferably, it is a medium size alkyl having 1 to 10 carbon atoms. Most preferably, it is a lower alkyl having 1 to 4 carbon atoms. The alkyl group may be substituted or unsubstituted. When substituted, the substituent group(s) is preferably one or more individually selected from cycloalkyl, aryl, heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, cyano, halo, carbonyl, thiocarbonyl, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, carboxy, nitro, silyl, amino and NR10R11.
R10 and R11 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl, sulfonyl, trihalomethanesulfonyl and, combined, a five-member or six-member heteroalicyclic ring.
A xe2x80x9ccycloalkylxe2x80x9d group refers to an all-carbon monocyclic or fused ring (i.e., rings which share an adjacent pair of carbon atoms) group wherein one of more of the rings does not have a completely conjugated pi-electron system. Examples, without limitation, of cycloalkyl groups are cyclopropane, cyclobutane, cyclopentane, cyclopentene, cyclohexane, cyclohexadiene, cycloheptane and, cycloheptatriene. A cycloalkyl group may be substituted or unsubstituted. When substituted, the substituent group(s) is preferably one or more individually selected from alkyl, aryl, heteroaryl, heteroalycyclic, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, cyano, halo, carbonyl, thiocarbonyl, carboxy, O-carbamyl, N-carbamyl, C-amido, N-amido, nitro, amino and NR10R11, with R10 and R11 being as defined above.
An xe2x80x9calkenylxe2x80x9d group refers to an alkyl group, as defined herein, consisting of at least two carbon atoms and at least one carbon-carbon double bond.
An xe2x80x9calkynylxe2x80x9d group refers to an alkyl group, as defined herein, consisting of at least two carbon atoms and at least one carbon-carbon triple bond.
An xe2x80x9carylxe2x80x9d group refers to an all-carbon monocyclic or fused-ring polycyclic (i.e., rings which share adjacent pairs of carbon atoms) groups having a completely conjugated pi-electron system. Examples, without limitation, of aryl groups are phenyl, naphthalenyl and anthracenyl. The aryl group may be substituted or unsubstituted. When substituted, the substituted group(s) is preferably one or more selected from halo, trihalomethyl, alkyl, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, cyano, nitro, carbonyl, thiocarbonyl, carboxy, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, sulfinyl, sulfonyl, amino and NR10R11 wherein R10 and R11 are previously defined herein.
As used herein, a xe2x80x9cheteroarylxe2x80x9d group refers to a monocyclic or fused ring (i.e., rings which share an adjacent pair of atoms) group having in the ring(s) one or more atoms selected from the group consisting of nitrogen, oxygen and sulfur and, in addition, having a completely conjugated pi-electron system. Examples, without limitation, of heteroaryl groups are pyrrole, furan, thiophene, imidazole, oxazole, thiazole, pyrazole, pyridine, pyrimidine, quinoline, isoquinoline, purine and carbazole. The heteroaryl group may be substituted or unsubstituted. When substituted, the substituted group(s) is preferably one or more selected from alkyl, cycloalkyl, halo, trihalomethyl, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, cyano, nitro, carbonyl, thiocarbonyl, sulfonamido, carboxy, sulfinyl, sulfonyl, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, amino and NR10R11 where R10 and R11 are previously defined herein.
A xe2x80x9cheteroalicyclicxe2x80x9d group refers to a monocyclic or fused ring group having in the ring(s) one or more atoms selected from the group consisting of nitrogen, oxygen and sulfur. The rings may also have one or more double bonds. However, the rings do not have a completely conjugated pi-electron system. The heteroalicyclic ring may be substituted or unsubstituted. When substituted, the substituted group(s) is preferably one or more selected from alkyl, cycloaklyl, halo, trihalomethyl, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, cyano, nitro, carbonyl, thiocarbonyl, carboxy, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, sulfinyl, sulfonyl, C-amido, N-amido, amino and NR10R11 where R10 and R11 are previously defined herein.
A xe2x80x9chydroxyxe2x80x9d group refers to an xe2x80x94OH group.
An xe2x80x9calkoxyxe2x80x9d group refers to both an xe2x80x94O-alkyl and an xe2x80x94O-cycloalkyl group, as defined herein.
An xe2x80x9caryloxyxe2x80x9d group refers to both an xe2x80x94O-aryl and an xe2x80x94O-heteroaryl group, as defined herein.
A xe2x80x9cthiohydroxyxe2x80x9d group refers to an xe2x80x94SH group.
A xe2x80x9cthioalkoxyxe2x80x9d group refers to both an S-alkyl and an xe2x80x94S-cycloalkyl group, as defined herein.
A xe2x80x9cthioaryloxyxe2x80x9d group refers to both an xe2x80x94S-aryl and an xe2x80x94S-heteroaryl group, as defined herein.
A xe2x80x9ccarbonylxe2x80x9d group refers to a xe2x80x94C(xe2x95x90O)xe2x80x94Rxe2x80x3 group, where Rxe2x80x3 is selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, heteroaryl (bonded through a ring carbon) and heteroalicyclic (bonded through a ring carbon), as defined herein.
An xe2x80x9caldehydexe2x80x9d group refers to a carbonyl group where Rxe2x80x3 is hydrogen.
A xe2x80x9cthiocarbonylxe2x80x9d group refers to a xe2x80x94C(xe2x95x90S)xe2x80x94Rxe2x80x3 group, with Rxe2x80x3 as defined herein.
A xe2x80x9ccarboxylxe2x80x9d group refers to a xe2x80x94C(xe2x95x90O)Oxe2x80x94Rxe2x80x3 groups, with Rxe2x80x3 as defined herein.
A xe2x80x9ccarboxylic acidxe2x80x9d group refers to a carboxyl group in which Rxe2x80x3 is hydrogen.
A xe2x80x9chaloxe2x80x9d group refers to fluorine, chlorine, bromine or iodine.
A xe2x80x9ctrihalomethylxe2x80x9d group refers to a xe2x80x94CX3 group wherein X is a halo group as defined herein.
A xe2x80x9ctrihalomethanesulfonylxe2x80x9d group refers to a X3CS(xe2x95x90O)2xe2x80x94 groups with X as defined above.
A xe2x80x9ccyanoxe2x80x9d group refers to a xe2x80x94Cxe2x95x90N group.
A xe2x80x9csulfinylxe2x80x9d group refers to a xe2x80x94S(xe2x95x90O)xe2x80x94Rxe2x80x3 group, with Rxe2x80x3 as defined herein.
A xe2x80x9csulfonylxe2x80x9d group refers to a xe2x80x94S(xe2x95x90O)2Rxe2x80x3 group, with Rxe2x80x3 as defined herein.
A xe2x80x9csulfonamidoxe2x80x9d group refers to a xe2x80x94S(xe2x95x90O)2NR10R11, with R10 and R11 as defined herein.
An xe2x80x9cO-carbamylxe2x80x9d group refers to a xe2x80x94OC(xe2x95x90O)NR10R11 group with R10 and R11 as defined herein.
An xe2x80x9cN-carbamylxe2x80x9d group refers to a R11OC(xe2x95x90O)NR10xe2x80x94 group, with R10 and R11 as defined herein.
An xe2x80x9cO-thiocarbamylxe2x80x9d group refers to a xe2x80x94OC(xe2x95x90S)NR10R11 group with R10 and R11 as defined herein.
An xe2x80x9cN-thiocarbamylxe2x80x9d group refers to a R11OC(xe2x95x90S)NR10xe2x80x94 group, with R10 and R11 as defined herein.
An xe2x80x9caminoxe2x80x9d group refers to an xe2x80x94NR10R11 group wherein R10 and R11 are both hydrogen.
A xe2x80x9cC-amidoxe2x80x9d group refers to a xe2x80x94C(xe2x95x90O)NR10R11 group with R10 and R11 as defined herein.
An xe2x80x9cN-amidoxe2x80x9d group refers to a R11C(xe2x95x90O)NR10xe2x80x94 group, with R10 and R11 as defined herein.
A xe2x80x9cquaternary ammoniumxe2x80x9d group refers to a xe2x80x94+NHR10R11 group wherein R10 and R11 are independently selected from the group consisting of alkyl, cycloalkyl, aryl, and heteroaryl.
A xe2x80x9cureidoxe2x80x9d group refers to a xe2x80x94NR10C(xe2x95x90O)NR11R12 group, with R10 and R11 as defined herein and R12 defined the same as R10 and R11.
A xe2x80x9cguanidinoxe2x80x9d group refers to a xe2x80x94R10NC(xe2x95x90N)NR11R12 group, with R10, R11 and R12 as defined herein.
A xe2x80x9cguanylxe2x80x9d group refers to a R10R11NC(xe2x95x90N)xe2x80x94 group, with R10 and R11 as defined herein.
A xe2x80x9cnitroxe2x80x9d group refers to a xe2x80x94NO2 group.
A xe2x80x9ccyanoxe2x80x9d group refers to a xe2x80x94Cxe2x89xa1N group.
A xe2x80x9csilylxe2x80x9d group refers to a xe2x80x94Si(Rxe2x80x3)3, with Rxe2x80x3 as defined herein.
A xe2x80x9cphosphonylxe2x80x9d group refers to a xe2x80x94OP(xe2x95x90O)2ORxe2x80x3, with Rxe2x80x3 as defined herein.
A xe2x80x9cheteroarylidenexe2x80x9d moiety refers to a heteroaryl-Cxe2x95x90Cxe2x80x94 group, with heteroaryl as defined herein.
xe2x80x9cMorpholinoxe2x80x9d refers to a group having the structure: 
xe2x80x9cPiperidinylxe2x80x9d refers to a group having the structure: 
xe2x80x9cPiperazinylxe2x80x9d refers to a group having the structure: 
xe2x80x9cTetrazoloxe2x80x9d refers to a group having the structure: 
B. Preferred Structural Features.
Preferred structural features of this invention are those in which:
J is selected from the group consisting of nitrogen and oxygen;
K, L and M are carbon;
R8 is selected from the group consisting of hydrogen, alkyl, halo, cyano, carboxyl, a five-member cycloalkyl, aryl, heteroaryl or heteoalicyclic ring fused to two adjacent atoms of the Q ring; and a six-member cycloalky, aryl, heteroaryl, or heteroalicyclic ring fused to two adjacent atoms of the Q ring;
alk1 is selected from the group consisting of CH2 and CHxe2x95x90CH;
n is 0, 1, 2 or 3; and,
Z is selected from the group consisting of hydroxy, alkoxy, amino, carboxyl, O-carbamyl, N-Carbamyl, C-amido, N-amido, morpholino, piperazinyl, tetrazolo, sulfonyl, sulfonamido, ureido, guanidinyl, and phosphonyl.
Additional preferred structural features of the compounds of this invention include those in which:
A, B, D, E, K, L and M are carbon;
R1 is hydrogen;
R8 is selected from the group consisting of hydrogen, alkyl, and a six-member cycloalky, aryl, heteroaryl, or heteroalicyclic ring fused to two adjacent atoms of the Q ring;
alk1 is CH2;
n is 1, 2 or 3;
Z is carboxyl;
R7 is hydrogen; and,
R3, R4, R5 and R6 are independently selected from the group comprising hydrogen, alkyl, trihaloalkyl, alkoxy, halo, carboxyl, nitro, amino, sulfonamido, and xe2x80x94NR10R11;
wherein
R10 and R11 are independently selected from the group comprising hydrogen, alkyl, carbonyl, carboxyl and sulfonyl.
Further preferred structural features of the compounds of this invention are those in which:
J and L are nitrogen;
R7 is selected from the group consisting of unsubstituted lower alkyl, unsubstituted aryl, unsubstituted heteroaryl, unsubstituted heteroalicyclic, sulfonyl, unsubstituted lower alkoxy, trihalomethanesulfonyl, and aryl or heteroaryl substituted with one or more groups selected from the group consisting of halo, amino, hydroxy, cyano, unsubstituted lower alkyl, unsubstituted lower alkoxy, carboxyl, S-sulfonamido, lower alkyl substituted with one or more groups selected from the group consisting of halo, hydroxy, amino or carboxyl and lower alkoxy substituted with one or more halo groups;
R8 is selected from the group consisting of unsubstituted lower alkyl, lower alkyl substited with one or more groups selected from the group consisting of halo, hydroxyl, unsubstituted lower alkoxy, amino or carboxyl, unsubstituted lower alkoxy, lower alkoxy substituted with one or more halo groups, unsubstituted aryl, unsubstituted heteroaryl, unsubstituted heteroalicyclic, aryl, heteroaryl or heteroalicyclic substituted with one or more groups selected from the group consisting of halogen, hydroxy, carboxy, nitro, cyano, amino, S-sulfonamido, unsubstituted lower alkoxy, lower alkoxy substituted with one or more halogens, unsubstituted lower alkyl or lower alkyl substituted with one or more groups selected from the group consisting of halogen, hydroxyl, amino or carboxyl;
R3, R4, R5 and R6 are independently selected from the groups consisting of hydrogen, halogen, nitro, amino, cyano, S-sulfonamido, carboxyl, trihalo-methyl, unsubstituted lower alkyl, lower alkyl substituted with one or more groups selected from the group consisting of halogen, hydroxyl, carboxyl, unsubstituted lower alkoxy and lower alkoxy substituted with one or more halo groups.
The chemical formulae referred to herein may exhibit the phenomena of tautomerism and structural isomerism. For example, the compounds described herein may adopt a cis or trans configuration about the double bond connecting the indolinone moiety to the heteroaryl moiety or they may be a mixture of cis and trans. This invention encompasses any tautomeric or structural isomeric form and mixtures thereof which possess the ability to modulate RTK, CTK and/or STK activity and is not limited to any one tautomeric or structural isomeric form.
As used herein, the term xe2x80x9ccisxe2x80x9d refers to the structural configuration wherein the heteroaryl group xe2x80x9cQxe2x80x9d is on the same side of the double bond connecting it to the 2-indolinone ring as the 2-oxygen group of the 2-indolinone.
As used herein, the term xe2x80x9ctransxe2x80x9d refers to the structural configuration wherein the heteroaryl group xe2x80x9cQxe2x80x9d is on the opposite side of the double bond connecting it to the 2-indolinone ring from the 2-oxygen group of the 2-indolinone.
Some of the preferred compounds of the invention are selected from the group consisting of 3-[3-(2-carboxyethyl-4-methylpyrrol-2-methylidenyl]-2-indolinone, 3-(2-acetyl-3,4-dimethylpyrrol-5-methylidenyl)-2-indolinone, 3-[4-(2-methoxycarbonylethyl-3-methylpyrrol-2-methylidenyl]-2-indolinone, 3-(2,4-dimethyl-3-ethoxycarbonylpyrrol-5-methylidenyl)-2-indolinone, 3-[2-ethoxycarbonyl-3-(2-ethoxycarbonylethyl)-4-ethoxycarbonylmethyl)pyrrol-5-methylidenyl)-2-indolinone, 3-(2-carboxy-4-ethyl-3-methylpyrrol-5-methylidenyl)-2-indolinone, 3-(2-chloro-4-methoxycarbonyl-3-methoxycarbonylmethylpyrrol-5-methylidenyl)-2-indolinone, 3-(4-acetyl-2-ethoxycarbonyl-3-methylpyrrol-5-methylidenyl)-2-indolinone, 3-(4-ethoxycarbonyl-3-methylpyrrol-2-methylidenyl)-2-indolinone, 3-[4-(2-methoxycarbonylethyl)-3-methylpyrrol-2-methylidenyl]-5,6-dimethoxy-2-indolinone, 3-(2,4-dimethyl-3-ethoxycarbonylpyrrol-5-methylidenyl)-5-(4-methoxycarbonylbenzamido)-2-indolinone, 3-(2,4-dimethyl-3-ethoxycarbonylpyrrol-5-methylidenyl)-5-bromo-2-indolinone, 3-[4-(2-carboxyethyl)-3,5-dimethylpyrrol-2-methylidenyl]-2-indolinone, and 3-[4-(2-carboxyethyl)-3-methylpyrrol-2-methylidenyl]-2-indolinone
C. Methods of Synthesis and Combinatorial Libraries
In another aspect, the invention provides a combinatorial library of at least 10 indolinone compounds that can be formed by reacting an oxindole with an acyl compound. The oxindole has a structure set forth in formula 2
where
A, B, D and E are selected from the group consisting of carbon and nitrogen, it being understood that the nitrogen-containing 9-member bicyclic ring formed is one known in the chemical arts; it being further understood that when A, B, D, or E is nitrogen, R3, R4, R5 or R6, respectively, does not exist;
R1 is selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, hydroxy, alkoxy, carboxyl, C-amido and sulfonyl;
R3, R4, R5 and R6 are independently selected from the group consisting of hydrogen, alkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, sulfonamido, carbonyl, carboxyl, cyano, nitro, halo, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, amino and xe2x80x94NR10R11, wherein
R10 and R11 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl, sulfonyl and, combined, a five- or six-member heteroalicyclic ring containing at least one nitrogen;
R3 and R4, R4 and R5, or R4 and R5 may combine to form a six-member aryl or heteroaryl ring;
and where the acyl compound has the structure set forth in formula 3
xe2x80x83where
J is selected from the group consisting of oxygen, nitrogen and sulfur;
K, L and M are independently selected from the group consisting of carbon, nitrogen, oxygen and sulfur such that the five-member heteroaryl ring formed is one known in the chemical arts, it being understood that when K, L and M are nitrogen, sulfur or oxygen, R8 or -(alk1)nZ cannot be covalently bonded to that atom;
when J is nitrogen, R7 is selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, hydroxy, alkoxy, aryloxy, carbonyl, carboxyl, C-amido, guanyl and sulfonyl and when J is oxygen or sulfur, R7 does not exist and there is no bond;
R8 is selected from the group consisting of hydrogen, alkyl, trihaloalkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, thiohydroxy, thioalkoxy, thioaryloxy, sulfinyl, sulfonyl, sulfonamido, carbonyl, carboxyl, cyano, nitro, halo, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, amino, xe2x80x94NR10R11, trihalomethyl, a five member cycloalkyl, aryl, heteroaryl or heteroalicyclic ring fused to two adjacent atoms of the five-membered ring; and a six-member cycloalky, aryl, heteroaryl, or heteroalicyclic ring fused to two adjacent atoms of the five-membered ring;
R10 and R11 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, carbonyl, sulfonyl and, combined, a five- or six-member heteroalicyclic ring containing at least one nitrogen;
alk1 is selected from the group consisting of optionally substituted methylene (xe2x80x94CRRxe2x80x2xe2x80x94), optionally substituted ethylene (xe2x80x94C(R)xe2x95x90C(Rxe2x80x2)xe2x80x94) and acetylene (xe2x80x94Cxe2x89xa1Cxe2x80x94);
R and Rxe2x80x2 are independently selected from the group consisting of hydrogen, alkyl, cycloalkyl, aryl, alkoxy, thioalkoxy, aryloxy and halo;
n is 0 to 10, inclusive; and
Z is a polar group.
The oxindole used in the combinatorial library is preferably selected from the group consisting of the indole compounds listed in Table 1 and the acyl compound used in the combinatorial library is preferably selected from the group consisting of the acyl compounds listed in Table 2.
A xe2x80x9ccombinatorial libraryxe2x80x9d refers to all the compounds formed by the reaction of each compound of one dimension with a compound in each of the other dimensions in a multi-dimensional array of compounds. In the context of the present invention, the array is two dimensional and one dimension represents all the oxindoles of the invention and the second dimension represents all the acyl compounds of the invention. Each oxindole may be reacted with each and every acyl compound in order to form an indolinone compound. All indolinone compounds formed in this way are within the scope of the present invention. Also within the scope of the present invention are smaller combinatorial libraries formed by the reaction of some of the oxindoles with all of the acyl compounds, all of the oxindoles with some of the acyl compounds, or some of the oxindoles with some of the acyl compounds.
By xe2x80x9cacyl compoundxe2x80x9d it is meant a compound of the formula Rxe2x80x94C(O)xe2x80x94Rxe2x80x2, where R and Rxe2x80x2 can be independently organic groups or hydrogen. Thus, aldehydes and ketones are examples of acyl compounds.
In order to form the combinatorial library of the present invention, all acyl compounds corresponding to the acyl portion of the compounds disclosed herein and all oxindole compounds corresponding to the indole portion of the compounds disclosed herein may be used in different combinations listed above.
Another aspect of the invention provides for a method for synthesizing an indolinone compound of formula 1, as described herein, comprising the step of reacting a first reactant with a second reactant in a solvent and in the presence of a base at elevated temperatures, where the first reactant is an oxindole having the structure set forth in formula 2 and the second reactant is an acyl compound, having a structure set forth in formula 3, as those formulae are described herein.
The first reactant is preferably an oxindole selected from the group consisting of the indole compounds listed in Table 1 and the second reactant is preferably an acyl compound selected from the group consisting of the acyl compounds listed in Table 2.
To synthesize the compounds of the invention a base may be used. The base is preferably a nitrogen base or an inorganic base. xe2x80x9cNitrogen basesxe2x80x9d are commonly used in the art and are selected from acyclic and cyclic amines. Examples of nitrogen bases include, but are not limited to, ammonia, methylamine, trimethylamine, triethylamine, aniline, 1,8-diazabicyclo[5.4.0]undec-7-ene, diisopropylethylamine, pyrrolidine, and piperidine. xe2x80x9cInorganic basesxe2x80x9d are bases that do not contain any carbon atoms. Examples of inorganic bases include, but are not limited to, hydroxide, phosphate, bisulfate, hydrosulfide, and amide anions. Those skilled in the art know which nitrogen base or inorganic base would match the requirements of the reaction conditions. In certain embodiments of the invention, the base used may be pyrrolidine or piperidine. In other embodiments the base may be the hydroxide anion, preferably used as its sodium or potassium salt.
The synthesis of the compounds of the invention takes place in a solvent. The solvent of the reaction is preferably a protic solvent or an aprotic solevent. xe2x80x9cProtic solventsxe2x80x9d are those that are capable of donating a proton to a solute. Examples of protic solvents include, but are not limited to, alcohols and water. xe2x80x9cAprotic solventsxe2x80x9d are those solvents that, under normal reaction conditions, do not donate a proton to a solute. Typical organic solvents, such as hexane, toluene, benzene, methylene chloride, dimethylformamide, chloroform, tetrahydrofuran, are some of the examples of aprotic solvents. Other aprotic solvents are also within the scope of used by the present invention. In some preferred embodiments, the solvent of the reaction is an alcohol, which may preferably be isopropanol or most preferably ethanol. Water is another preferred protic solvent. Dimethylformamide, known in the chemistry art as DMF, is a preferred aprotic solvent.
The synthetic method of the invention calls for the reaction to take place at elevated temperatures which are temperatures that are greater than room temperature. More preferably, the elevated temperature is preferably about 30-150xc2x0 C., more preferably is about 80-100xc2x0 C., and most preferably is about 80-90xc2x0 C., which is about the temperature at which ethanol boils (i.e., the boiling point of ethanol). By xe2x80x9caboutxe2x80x9d a certain temperature it is meant that the temperature range is preferably within 10xc2x0 C. of the listed temperature, more preferably within 5xc2x0 C. of the listed temperature, and most preferably within 2xc2x0 C. of the listed temperature. Therefore, by way of example, by xe2x80x9cabout 80xc2x0 C.xe2x80x9d it is meant that the temperature range is preferably 80xc2x110xc2x0 C., more preferably 80xc2x15xc2x0 C., and most preferably 80xc2x12xc2x0 C.
The synthetic method of the invention may be accompanied by the step of screening a library for a compound of the desired activity and structurexe2x80x94thus, providing a method of synthesis of a compound by first screening for a compound having the desired properties and then chemically synthesizing that compound.
In yet another embodiment, this invention relates to a method for the modulation of the catalytic activity of PKs by contacting a PK with a compound of this invention or a physiologically acceptable salt or prodrug thereof.
By xe2x80x9cPKxe2x80x9d is meant RTK, CTK and STK; i.e., the modulation of RTK, CTK and STK catalyzed signaling processes are contemplated by this invention.
The term xe2x80x9cmethodxe2x80x9d refers to manners, means, techniques and procedures for accomplishing a given task including, but not limited to, those manners, means, techniques and procedures either known to, or readily developed from known manners, means, techniques and procedures by, practitioners of the chemical, pharmacological, biological, biochemical and medical arts.
As used herein, the term xe2x80x9cmodulationxe2x80x9d or xe2x80x9cmodulatingxe2x80x9d refers to the alteration of the catalytic activity of RTKs, CTKs and STKs. In particular, modulating refers to the activation of the catalytic activity of RTKs, CTKs and STKs, preferably the activation or inhibition of the catalytic activity of RTKs, CTKs and STKs, depending on the concentration of the compound or salt to which the RTK, CTK or STK is exposed or, more preferably, the inhibition of the catalytic activity of RTKs, CTKs and STKs.
The term xe2x80x9ccatalytic activityxe2x80x9d as used herein refers to the rate of phosphorylation of tyrosine under the influence, direct or indirect of RTKs and/or CTKs or the phosphorylation of serine and threonine under the influence, direct or indirect, of STKs.
The term xe2x80x9ccontactingxe2x80x9d as used herein refers to bringing a compound of this invention and a target PK together in such a manner that the compound can affect the catalytic activity of the PK, either directly; i.e., by interacting with the kinase itself, or indirectly; i.e., by interacting with another molecule on which the catalytic activity of the kinase is dependent. Such xe2x80x9ccontactingxe2x80x9d can be accomplished in a test tube, a petri dish or the like. In a test tube, contacting may involve only a compound and a PK of interest or it may involve whole cells. Cell may also be maintained or grown in cell culture dishes and contacted with the compound in that environment. In this context, the ability of a particular compound to affect a PK related disorder; i.e., the IC50 of the compound, defined below, can be determined before the compounds are used in vivo with more complex living organisms is attempted. For cells outside the organism, multiple methods exist, and are well-known to those skilled in the arts, to get the PKs in contact with the compounds including, but not limited to, direct cell microinjection and numerous transmembrane carrier techniques.
RTK mediated signal transduction is initiated by extracellular interaction with a specific growth factor (ligand), followed by receptor dimerization, transient stimulation of the intrinsic protein tyrosine kinase activity and phosphorylation. Binding sites are thereby created for intracellular signal transduction molecules and lead to the formation of complexes with a spectrum of cytoplasmic signaling molecules that facilitate the appropriate cellular response (e.g., cell division, metabolic effects to the extracellular micro-environment). See, Schlessinger and Ullrich, 1992, Neuron 9:303-391.
It has been shown that tyrosine phosphorylation sites in growth factor receptors function as high-affinity binding sites for SH2 (src homology) domains of signaling molecules. Fantl et al., 1992, Cell 69:413-423; Songyang et al., 1994, Mol. Cell. Biol. 14:2777-2785); Songyang et al., 1993, Cell 72:767-778; and Koch et al., 1991, Science 252:668-678. Several intracellular substrate proteins that associate with RTKs have been identified. They may be divided into two principal groups: (1) substrates which have a catalytic domain; and (2) substrates which lack such domain but serve as adapters and associate with catalytically active molecules. Songyang et al., 1993, Cell 72:767-778. The specificity of the interactions between receptors and SH2 domains of their substrates is determined by the amino acid residues immediately surrounding the phosphorylated tyrosine residue. Differences in the binding affinities between SH2 domains and the amino acid sequences surrounding the phosphotyrosine residues on particular receptors are consistent with the observed differences in their substrate phosphorylation profiles. Songyang et al., 1993, Cell 72:767-778. These observations suggest that the function of each RTK is determined not only by its pattern of expression and ligand availability but also by the array of downstream signal transduction pathways that are activated by a particular receptor. Thus, phosphorylation provides an important regulatory step which determines the selectivity of signaling pathways recruited by specific growth factor receptors, as well as differentiation factor receptors.
STKs, being primarily cytosolic, affect the internal biochemistry of the cell often as a down-line response to a PTK event. STKs have been implicated in the signaling process which initiates DNA synthesis and subsequent mitosis leading to cell proliferation.
Thus, PK signal transduction results in, among other responses, cell proliferation, differentiation, growth and metabolism. Abnormal cell proliferation may result in a wide array of disorders and diseases, including the development of neoplasia such as carcinoma, sarcoma, leukemia, glioblastoma, hemangioma, psoriasis, arteriosclerosis, arthritis and diabetic retinopathy (or other disorders related to uncontrolled angiogenesis and/or vasculogenesis).
A precise understanding of the mechanism by which the compounds of this invention inhibit PKs is not required in order to practice the present invention. However, while not hereby being bound to any particular mechanism or theory, it is believed that the compounds interact with the amino acids of the catalytic region of PKs. PKs typically possess a bi-lobate structure wherein ATP appears to bind in the cleft between the two lobes in a region where the amino acids are conserved among PKs. Inhibitors of PKs are believed to bind by non-covalent interactions such as hydrogen bonding, van der Waals forces and ionic interactions in the same general region where the aforesaid ATP binds to the PKs. More specifically, it is thought that the 2-indolinone component of the compounds of this invention binds in the general space normally occupied by the adenine ring of ATP. Specificity of a particular molecule for a particular PK could arise as the result of additional interactions between the various substituents on the 2-indolinone core with amino acid domains specific to particular PKs. Thus, different indolinone substituents may contribute to preferential binding to particular PKs. The ability to select those compounds active at different ATP (or other nucleotide) binding sites makes the compounds useful for targeting any protein with such a site; i.e., not only PKs but protein phosphatases as well. The compounds disclosed herein thus have utility for in vitro assays on such proteins and for in vivo therapeutic effects through such proteins.
In another aspect, the protein kinase, the catalytic activity of which is modulated by contact with a compound of this invention, is a protein tyrosine kinase, more particularly, a receptor protein tyrosine kinase. Among the receptor protein tyrosine kinases whose catalytic activity can be modulated with a compound of this invention, or salt thereof, are, without limitation, EGF, HER2, HER3, HER4, IR, IGF-1R, IRR, PDGFRxcex1, PDGFRxcex2, CSFIR, C-Kit, C-fms, Flk-1R, Flk4, KDR/Flk-1, Flt-1, FGFR-1R, FGFR-2R, FGFR-3R and FGFR-4R.
The protein tyrosine kinase whose catalytic activity is modulated by contact with a compound of this invention, or a salt or a prodrug thereof, can also be a non-receptor or cellular protein tyrosine kinase (CTK). Thus, the catalytic activity of CTKs such as, without limitation, Src, Frk, Btk, Csk, Abl, ZAP70, Fes/Fps, Fak, Jak, Ack, Yes, Fyn, Lyn, Lck, Blk, Hck, Fgr and Yrk, may be modulated by contact with a compound or salt of this invention.
Still another group of PKs which may have their catalytic activity modulated by contact with a compound of this invention are the serine-threonine protein kinases such as, without limitation, CDK2 and Raf.
In another aspect, this invention relates to a method for treating or preventing a PK related disorder by administering a therapeutically effective amount of a pharmacological composition of this compound of this invention or a salt or a prodrug thereof to an organism.
As used herein, xe2x80x9cPK related disorder,xe2x80x9d xe2x80x9cPK driven disorder,xe2x80x9d and xe2x80x9cabnormal PK activityxe2x80x9d all refer to a condition characterized by inappropriate; i.e., under or, more commonly, over, PK catalytic activity, where the particular PK be an RTK, a CTK or an STK. Inappropriate catalytic activity can arise as the result of either: (1) PK expression in cells which normally do not express PKs; (2) increased PK expression leading to unwanted cell proliferation, differentiation and/or growth; or, (3) decreased PK expression leading to unwanted reductions in cell proliferation, differentiation and/or growth. Over-activity of PKs refers to either amplification of the gene encoding a particular PK or production of a level of PK activity which can correlate with a cell proliferation, differentiation and/or growth disorder (that is, as the level of the PK increases, the severity of one or more of the symptoms of the cellular disorder increases). Underactivity is, of course, the converse, wherein the severity of one or more symptoms of a cellular disorder increase as the level of the PK decreases.
As used herein, the terms xe2x80x9cpreventxe2x80x9d, xe2x80x9cpreventingxe2x80x9d and xe2x80x9cpreventionxe2x80x9d refer to a method for barring an organism from in the first place acquiring an PK mediated cellular disorder.
As used herein, the terms xe2x80x9ctreatxe2x80x9d, xe2x80x9ctreatingxe2x80x9d and xe2x80x9ctreatmentxe2x80x9d refer to a method of alleviating or abrogating the PK mediated cellular disorder and/or its attendant symptoms. With regard particularly to cancer, these terms simply mean that the life expectancy of an individual affected with a cancer will be increased or that one or more of the symptoms of the disease will be reduced.
The term xe2x80x9corganismxe2x80x9d refers to any living entity comprised of at least one cell. A living organism can be as simple as, for example, a single eukariotic cell or as complex as a mammal, including a human being.
The term xe2x80x9ctherapeutically effective amountxe2x80x9d as used herein refers to that amount of the compound being administered which will relieve to some extent one or more of the symptoms of the disorder being treated. In reference to the treatment of cancer, a therapeutically effective amount refers to that amount which has the effect of (1) reducing the size of the tumor; (2) inhibiting (that is, slowing to some extent, preferably stopping) tumor metastasis; (3) inhibiting to some extent (that is slowing to some extent, preferably stopping) tumor growth; and/or, (4) relieving to some extent (or preferably eliminating) one or more symptoms associated with the cancer.
This invention is therefore directed to compounds which modulate PK signal transduction by affecting the enzymatic activity of the RTKs, CTKs and/or STKs and thereby interfering with the signal transduced by such proteins. More particularly, the present invention is directed to compounds which modulate the RTK, CTK and/or STK mediated signal transduction pathways as a therapeutic approach to cure many kinds of solid tumors, including but not limited to carcinoma, sarcomas including Kaposi""s sarcoma, leukemia, erythroblastoma, glioblastoma, meningioma, astrocytoma, melanoma and myoblastoma. Indications may include, but are not limited to brain cancers, bladder cancers, ovarian cancers, gastric cancers, pancreas cancers, colon cancers, blood cancers, lung cancers, bone cancers and leukemias.
Further examples, without limitation, of the types of disorders related to unregulated PK activity that the compounds described herein may be useful in preventing, treating and studying, are cell proliferative disorders, fibrotic disorders and metabolic disorders.
Cell proliferative disorders, which may be prevented, treated or further studied by the present invention include cancers, blood vessel proliferative disorders and mesangial cell proliferative disorders.
Blood vessel proliferative disorders refer to angiogenic and vasculogenic disorders generally resulting in abnormal proliferation of blood vessels. The formation and spreading of blood vessels, or vasculogenesis and angiogenesis, respectively, play important roles in a variety of physiological processes such as embryonic development, corpus luteum formation, wound healing and organ regeneration. They also play a pivotal role in cancer development. Other examples of blood vessel proliferation disorders include arthritis, where new capillary blood vessels invade the joint and destroy cartilage, and ocular diseases, like diabetic retinopathy, where new capillaries in the retina invade the vitreous, bleed and cause blindness. Conversely, disorders related to the shrinkage, contraction or closing of blood vessels, such as restenosis, are also implicated.
Fibrotic disorders refer to the abnormal formation of extracellular matrices. Examples of fibrotic disorders include hepatic cirrhosis and mesangial cell proliferative disorders. Hepatic cirrhosis is characterized by the increase in extracellular matrix constituents resulting in the formation of a hepatic scar. Hepatic cirrhosis can cause diseases such as cirrhosis of the liver. An increased extracellular matrix resulting in a hepatic scar can also be caused by viral infection such as hepatitis. Lipocytes appear to play a major role in hepatic cirrhosis. Other fibrotic disorders implicated include atherosclerosis.
Mesangial cell proliferative disorders refer to disorders brought about by abnormal proliferation of mesangial cells. Mesangial proliferative disorders include various human renal diseases, such as glomerulonephritis, diabetic nephropathy, malignant nephrosclerosis, thrombotic microangiopathy syndromes, transplant rejection, and glomerulopathies. The PDGF-R has been implicated in the maintenance of mesangial cell proliferation. Floege et al., 1993, Kidney International 43:47S-54S.
As noted previously, PKs have been associated with such cell proliferative disorders. For example, some members of the RTK family have been associated with the development of cancer. Some of these receptors, like the EGFR (Tuzi et al., 1991, Br. J. Cancer 63:227-233; Torp et al., 1992, APMIS 100:713-719) HER2/neu (Slamon et al., 1989, Science 244:707-712) and the PDGF-R (Kumabe et al., 1992, Oncogene, 7:627-633) are over-expressed in many tumors and/or persistently activated by autocrine loops. In fact, in the most common and severe cancers these receptor over-expressions (Akbasak and Suner-Akbasak et al., 1992, J. Neurol. Sci., 111:119-133; Dickson et al., 1992, Cancer Treatment Res. 61:249-273; Korc et al., 1992, J. Clin. Invest. 90:1352-1360) and autocrine loops (Lee and Donoghue, 1992, J. Cell. Biol., 118:1057-1070; Korc et al., supra; Akbasak and Suner-Akbasak et al., supra) have been demonstrated. For example, the EGFR receptor has been associated with squamous cell carcinoma, astrocytoma, glioblastoma, head and neck cancer, lung cancer and bladder cancer. HER2 has been associated with breast, ovarian, gastric, lung, pancreas and bladder cancer. PDGFR has been associated with glioblastoma, lung, ovarian, melanoma and prostate. The RTK c-met has been generally associated with hepatocarcinogenesis and thus hepatocellular carcinoma. C-met has been linked to malignant tumor formation. More specifically, the RTK c-met has been associated with, among other cancers, colorectal, thyroid, pancreatic and gastric carcinoma, leukemia and lymphoma. Additionally, over-expression of the c-met gene has been detected in patients with Hodgkins disease, Burkitts disease, and the lymphoma cell line.
Flk has been associated with a broad spectrum of tumors including without limitation mammary, ovarian and lung tumors as well as gliomas such as glioblastoma.
IGF-IR, in addition to being implicated in nutritional support and in type-II diabetes, has also been associated with several types of cancers. For example, IGF-I has been implicated as an autocrine growth stimulator for several tumor types, e.g. human breast cancer carcinoma cells (Arteaga et al., 1989, J. Clin. Invest. 84:1418-1423) and small lung tumor cells (Macauley et al., 1990, Cancer Res., 50:2511-2517). In addition, IGF-I, integrally involved in the normal growth and differentiation of the nervous system, appears to be an autocrine stimulator of human gliomas. Sandberg-Nordqvist et al., 1993, Cancer Res. 53:2475-2478. The importance of the IGF-IR and its ligands in cell proliferation is further supported by the fact that many cell types in culture (fibroblasts, epithelial cells, smooth muscle cells, T-lymphocytes, myeloid cells, chondrocytes, osteoblasts, the stem cells of the bone marrow) are stimulated to grow by IGF-I. Goldring and Goldring, 1991, Eukaryotic Gene Expression, 1:301-326. In a series of recent publications, Baserga even suggests that IGF-IR plays a central role in the mechanisms of transformation and, as such, could be a preferred target for therapeutic interventions for a broad spectrum of human malignancies. Baserga, 1995, Cancer Res., 55:249-252; Baserga, 1994, Cell 79:927-930; Coppola et al., 1994, Mol. Cell. Biol., 14:4588-4595.
STKs have been implicated in many types of cancer including notably breast cancer (Cance, et al., Int. J. Cancer, 54:571-77 (1993)).
The association between abnormal PK activity and disease are not restricted to cancer, however. For example, RTKs have been associated with diseases such as psoriasis, diabetes mellitus, endometriosis, angiogenesis, atheromatous plaque development, Alzheimer""s disease, epidermal hyperproliferation and neurodegenerative diseases age-related macular degeneration, hemangiomas. For example, the EGF-R is indicated in corneal and dermal wound healing. Defects in the Insulin-R and the IGF-1R are indicated in type-II diabetes mellitus. A more complete correlation between specific RTKs and their therapeutic indications is set forth in Plowman et al., 1994, DNandP 7:334-339.
As noted previously, not only RTKs but CTKs as well including, but not limited to, src, abl, fps, yes, fyn, lyn, lck, blk, hck, fgr and yrk (reviewed by Bolen et al., 1992, FASEB J., 6:3403-3409) are involved in the proliferative and metabolic signal transduction pathway and thus were expected, and have been shown, to be involved in many PTK-mediated disorders to which the present invention is directed. For example, mutated src (v-src) has been demonstrated as an oncoprotein (pp60v-src) in chicken. Moreover, its cellular homolog, the proto-oncogene pp60c-src transmits oncogenic signals of many receptors. For example, over-expression of EGFR or HER2/neu in tumors leads to the constitutive activation of pp60cXsrc, which is characteristic for the malignant cell but absent from the normal cell. On the other hand, mice deficient in the expression of c-src exhibit an osteopetrotic phenotype, indicating a key participation of c-src in osteoclast function and a possible involvement in related disorders. Similarly, Zap70 is implicated in T-cell signaling.
STKs have been associated with inflamation, autoimmune disease, immunoresponses, and hyperproliferation disorders such as restinosis, fibrosis, psoriasis, osteoarthritis and rheumatoid arthritis.
PKs have also been implicated in embryo implantation and the compounds of this invention may provide an effective method of preventing embryo implantation.
Finally, both RTKs and CTKs are currently suspected as being involved in hyperimmune disorders.
A compound of the present invention, a prodrug thereof or its physiologically acceptable salt of either the salt or prodrug can be administered as such to a human patient or in pharmacological compositions where these are mixed with suitable carriers or excipient (s). Techniques for formulation and administration of drugs may be found in xe2x80x9cRemington""s Pharmaceutical Sciences,xe2x80x9d Mack Publishing Co., Easton, Pa., latest edition.
As used herein, xe2x80x9cadministerxe2x80x9d or xe2x80x9cadministrationxe2x80x9d refers to the delivery of a compound, salt or prodrug of the present invention or of a pharamacological composition containing a compound, salt or prodrug of this invention to an organism for the purpose of prevention or treatment of a PK-related disorder.
Suitable routes of administration may include, without limitation, oral, rectal, transmucosal or intestinal administration; or, intramuscular, subcutaneous, intramedullary, intrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal, or intraocular injections.
Alternately, one may administer the compound in a local rather than systemic manner, for example, via injection of the compound directly into a solid tumor, often in a depot or sustained release formulation.
Furthermore, one may administer the drug in a targeted drug delivery system, for example, in a liposome coated with tumor-specific antibody. The liposomes will be targeted to and taken up selectively by the tumor.
Pharmacological compositions of the present invention may be manufactured by processes well known in the art; e.g., by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or lyophilizing processes.
Pharmacological compositions for use in accordance with the present invention thus may be formulated in conventional manner using one or more physiologically acceptable carriers comprising excipients and auxiliaries which facilitate processing of the active compounds into preparations which can be used pharmaceutically. Proper formulation is dependent upon the route of administration chosen.
For injection, the compounds of the invention may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hanks""s solution, Ringer""s solution, or physiological saline buffer. For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.
For oral administration, the compounds can be formulated readily by combining the active compounds with pharmaceutically acceptable carriers well known in the art. Such carriers enable the compounds of the invention to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a patient to be treated. Pharmacological preparations for oral use can be made with the use of a solid excipient, optionally grinding the resulting mixture, and processing the mixture of granules, after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores. Suitable excipients are, in particular, fillers such as sugars, including lactose, sucrose, mannitol, or sorbitol; cellulose preparations such as, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, and/or polyvinylpyrrolidone (PVP). If desired, disintegrating agents may be added, such as the cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate.
Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures. Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses.
Pharmacological compositions which can be used orally include push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. The push-fit capsules can contain the active ingredients in admixture with filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In addition, stabilizers may be added. All formulations for oral administration should be in dosages suitable for such administration.
For buccal administration, the compositions may take the form of tablets or lozenges formulated in conventional manner.
For administration by inhalation, the compounds for use according to the present invention are conveniently delivered in the form of an aerosol spray presentation from pressurized packs or a nebulizer, with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichloro-tetrafluoroethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of, for example, gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of the compound and a suitable powder base such as lactose or starch.
The compounds may be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion. Formulations for injection may be presented in unit dosage form, e.g., in ampoules or in multi-dose containers, with an added preservative. The compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
Pharmacological compositions for parenteral administration include aqueous solutions of the active compounds in water soluble form. Additionally, suspensions of the active compounds may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds to allow for the preparation of highly concentrated solutions.
Alternatively, the active ingredient may be in powder form for constitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use.
The compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter or other glycerides.
In addition to the formulations described previously, the compounds may also be formulated as a depot preparation. Such long acting formulations may be administered by implantation (for example subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the compounds may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.
The pharmacological compositions herein also may comprise suitable solid or gel phase carriers or excipients. Examples of such carriers or excipients include but are not limited to calcium carbonate, calcium phosphate, various sugars, starches, cellulose derivatives, gelatin, and polymers such as polyethylene glycols.
Many of the PK modulating compounds of the invention may be provided as physiologically acceptable salts wherein the claimed compound may form the negatively or the positively charged species. Examples of salts in which the compound forms the positively charged moiety include, without limitation, quaternary ammonium (defined elsewhere herein), salts such as the hydrochloride, sulfate, carbonate, lactate, tartrate, maleate, succinate, etc. formed by the reaction of an amino group with the appropriate acid. Salts in which the compound forms the negatively charged species include, without limitation, the sodium, potassium, calcium and magnesium salts formed by the reaction of a carboxylic acid group in the molecule with the appropriate base (e.g. sodium hydroxide (NaOH), potassium hydroxide (KOH), Calcium hydroxide (Ca(OH)2), etc.).
Pharmacological compositions suitable for use in the present invention include compositions wherein the active ingredients are contained in an amount effective to achieve its intended purpose.
More specifically, a therapeutically effective amount means an amount of compound effective to prevent, alleviate or ameliorate symptoms of disease or prolong the survival of the subject being treated.
Determination of a therapeutically effective amount is well within the capability of those skilled in the art, especially in light of the detailed disclosure provided herein.
For any compound used in the methods of the invention, the therapeutically effective amount or dose can be estimated initially from cell culture assays. Then, the dosage can be formulated for use in animal models so as to achieve a circulating concentration range that includes the IC50 as determined in cell culture (i.e., the concentration of the test compound which achieves a half-maximal inhibition of the PK activity). Such information can then be used to more accurately determine useful doses in humans.
Toxicity and therapeutic efficacy of the compounds described herein can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the LD50 (the dose lethal to 50% of the population) and the ED50 (the dose therapeutically effective in 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index and it can be expressed as the ratio between LD50 and ED50. Compounds which exhibit high therapeutic indices are preferred. The data obtained from these cell culture assays and animal studies can be used in formulating a range of dosage for use in human. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized. The exact formulation, route of administration and dosage can be chosen by the individual physician in view of the patient""s condition. (See e.g., Fingl, et al., 1975, in xe2x80x9cThe Pharmacological Basis of Therapeuticsxe2x80x9d, Ch. 1 p.1).
Dosage amount and interval may be adjusted individually to provide plasma levels of the active moiety which are sufficient to maintain the kinase modulating effects, or minimal effective concentration (MEC). The MEC will vary for each compound but can be estimated from in vitro data; e.g., the concentration necessary to achieve 50-90% inhibition of the kinase using the assays described herein. Dosages necessary to achieve the MEC will depend on individual characteristics and route of administration. However, HPLC assays or bioassays can be used to determine plasma concentrations.
Dosage intervals can also be determined using MEC value. Compounds should be administered using a regimen which maintains plasma levels above the MEC for 10-90% of the time, preferably between 30-90% and most preferably between 50-90%.
In cases of local administration or selective uptake, the effective local concentration of the drug may not be related to plasma concentration.
The amount of composition administered will, of course, be dependent on the subject being treated, on the subject""s weight, the severity of the affliction, the manner of administration and the judgment of the prescribing physician.
The compositions may, if desired, be presented in a pack or dispenser device, such as an FDA approved kit, which may contain one or more unit dosage forms containing the active ingredient. The pack may for example comprise metal or plastic foil, such as a blister pack. The pack or dispenser device may be accompanied by instructions for administration. The pack or dispenser may also be accompanied by a notice associated with the container in a form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals, which notice is reflective of approval by the agency of the form of the compositions or human or veterinary administration. Such notice, for example, may be of the labeling approved by the U.S. Foods and Drug Administration for prescription drugs or of an approved product insert. Compositions comprising a compound of the invention formulated in a compatible pharmaceutical carrier may also be prepared, placed in an appropriate container, and labeled for treatment of an indicated condition. Suitable conditions indicated on the label may include treatment of a tumor, inhibition of angiogenesis, treatment of fibrosis, diabetes, and the like.
Table 1 shows a representative, but by no means exclusive, list of 2-indolinones which may be used in the synthesis of the compounds of this invention by reaction with any of the aldehydes of Table 2.
Table 2 shows a representative, but by no means exclusive, list of aldehydes which may be used in the synthesis of the compounds of this invention by reaction with any of the 2-indolinones of Table 1. Table 3 shows the results of biological assays of several of the preferred compounds of this invention. PDGFR, FLK-1R, EGFR HER2 and IGF-1R are discussed above. IC50 refers to that amount of the tested compound needed to effect a 50% change in the activity of the PTK in the test indicated with respect to a control in which no compound of this invention is present. With regard to the tests in the table, the 50% change being evaluated is a 50% inhibition of PTK activity over that of the control.